Signaling mechanisms regulating vascular smooth muscle mitogenesis are being clearly elucidated and exploited for potential therapeutic benefit. To date however, there are still no effective therapeutics designed to control dysregulated vascular smooth muscle proliferation during inflammatory pathologies. Our laboratory is focused on identifying and characterizing endogenous lipid-derived second messengers that inhibit pro-mitogenic signaling cascades regulated through protein kinase C (PKC) and phosphoinositide-3-kinase (PI3K). Using in vivo and in vitro model systems, we have demonstrated that interleukin-1-generated ether-linked diglycerides (ether-DG) inhibit smooth muscle cell mitogenesis. These novel phospholipid-derived second messengers mimic the effect of interleukin-1 to inhibit cellular proliferation. As ether-DGs are analogues of diacylglycerol (DAG), a co- factor for growth factor-stimulated PKC activation, we hypothesize that ether-DGs can competitively antagonize DAG-activated PKC as a mechanism to diminish smooth muscle mitogenesis. We now demonstrate that DAG analogues enhance PI3K activity. Thus, we also hypothesize that ether-DGs diminish cellular proliferation by competitively antagonizing DAG-stimulated PI3K activity. In Specific Aim 1, the biochemical mechanisms by which ether-DGs inhibit PKC delta and epsilon activation will be investigated in vascular smooth muscle cells. In Specific Aim 2, the role of ether-DGs to inhibit mitogenesis through PKC-dependent or -independent inhibition of P13K will be investigated. These studies will establish ether-DGs as potential therapeutics to limit abnormal vascular smooth muscle cell growth observed in atherosclerotic and restenotic lesions.